Naomasa Fukase

Influence of intra-operative joint gap on post-operative flexion angle in osteoarthritis patients undergoing posterior-stabilized total knee arthroplasty

Recently, we developed a new tensor for total knee arthroplasty (TKA) procedures enabling soft tissue balance assessment throughout the range of motion while reproducing post-operative joint alignment with the patello-femoral (PF) joint reduced and the tibiofemoral joint aligned. Using the tensor with a computer-assisted navigation system, we investigated the relationship between various intra-operative joint gap values and their post-operative flexion angles. An increased value during the extension to flexion gap and a decreased value during the flexion to deep flexion gap with PF joint reduced, not everted, showed an inverse correlation with post-operative knee flexion angle, not pre-operative flexion angle. In conclusion, understanding the characteristics of joint gap kinematics in posterior-stabilized TKA under physiological and reproducible joint conditions may enable the prediction of the post-operative flexion angle and help to determine the appropriate intra-operative joint gap.

PKD1 negatively regulates cell invasion, migration and proliferation ability of human osteosarcoma.

Osteosarcoma (OS) is a primary malignancy of the bone, with a tendency to metastasize early. Despite intensive chemotherapy and surgical resection, more than 30% of patients develop distant metastases, and the prognosis of patients with metastases is essentially poor. Members of the protein kinase D (PKD) family are serine/threonine kinases, and have been studied in various cancers. Among the three different isoforms of this family, PKD1 is one of the best understood for its role in human malignancies; however, its role in musculoskeletal tumors has not been studied. In the present study, we investigated the role of PKD1 in human OS. We first analyzed PKD1 mRNA expression in human musculoskeletal tumor tissue samples by quantitative real-time PCR. PKD1 expression in OS samples was significantly lower than that in benign schwannoma samples, and this was correlated with metastatic potential. In in vitro studies, overexpression of PKD1 by plasmid transfection decreased OS cell invasion, migration and proliferation, and significantly decreased matrix metalloproteinase (MMP)2 mRNA expression. Conversely, siRNA knockdown of PKD1 increased invasion, migration and proliferation of OS cells, and MMP2 expression was markedly increased. Furthermore, overexpression of PKD1 significantly reduced in vivo tumor growth of OS cells. These results demonstrated that low expression of PKD1 may contribute to increased cell invasion, migration and proliferation ability of human OS. Taken together, our findings strongly suggest that PKD1 may negatively regulate the malignant potential of human OS, and may be a therapeutic target for human OS in the clinical setting.

Reoxygenation using a novel CO2 therapy decreases the metastatic potential of osteosarcoma cells.

Osteosarcoma is the most common primary solid malignant bone tumor. Despite substantial improvements in surgery and chemotherapy, metastasis remains a major cause of fatal outcomes, and the molecular mechanisms of metastasis are still poorly understood. Hypoxia, which is common in malignant tumors including osteosarcoma, increases expressions of hypoxia inducible factor (HIF)-1α, matrix metalloproteinase (MMP)-2 and MMP-9, and can induce invasiveness. As we previously showed a novel transcutaneous CO2 application to decrease HIF-1α expression and induce apoptosis in malignant fibrous histiocytoma, we hypothesize that transcutaneous CO2 application could suppress metastatic potential of osteosarcoma by improving hypoxic conditions. Here, we examined the effects of transcutaneous CO2 application on apoptosis, and development of pulmonary metastasis using a highly metastatic osteosarcoma cell line, LM8. Transcutaneous CO2 application significantly decreased tumor growth and pulmonary metastasis in LM8 cells. Apoptotic activity increased, and intratumoral hypoxia was improved with decreased expressions of HIF-1α, MMP-2 and MMP-9, significantly, in the CO2-treated tumors. In conclusion, we found that transcutaneous CO2 application can induce tumor cell apoptosis and might suppress pulmonary metastasis by improvement of hypoxic conditions with decreased expressions of HIF-1α and MMPs in highly metastatic osteosarcoma cell. These findings strongly indicate that this novel transcutaneous CO2 therapy could be a therapeutic breakthrough for osteosarcoma patients.

Transcutaneous Application of Carbon Dioxide (CO2) Induces Mitochondrial Apoptosis in Human Malignant Fibrous Histiocytoma In Vivo

Mitochondria play an essential role in cellular energy metabolism and apoptosis. Previous studies have demonstrated that decreased mitochondrial biogenesis is associated with cancer progression. In mitochondrial biogenesis, peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) regulates the activities of multiple nuclear receptors and transcription factors involved in mitochondrial proliferation. Previously, we showed that overexpression of PGC-1α leads to mitochondrial proliferation and induces apoptosis in human malignant fibrous histiocytoma (MFH) cells in vitro. We also demonstrated that transcutaneous application of carbon dioxide (CO2) to rat skeletal muscle induces PGC-1α expression and causes an increase in mitochondrial proliferation. In this study, we utilized a murine model of human MFH to determine the effect of transcutaneous CO2 exposure on PGC-1α expression, mitochondrial proliferation and cellular apoptosis. PGC-1α expression was evaluated by quantitative real-time PCR, while mitochondrial proliferation was assessed by immunofluorescence staining and the relative copy number of mitochondrial DNA (mtDNA) was assessed by real-time PCR. Immunofluorescence staining and DNA fragmentation assays were used to examine mitochondrial apoptosis. We also evaluated the expression of mitochondrial apoptosis related proteins, such as caspases, cytochorome c and Bax, by immunoblot analysis. We show that transcutaneous application of CO2 induces PGC-1α expression, and increases mitochondrial proliferation and apoptosis of tumor cells, significantly reducing tumor volume. Proteins involved in the mitochondrial apoptotic cascade, including caspase 3 and caspase 9, were elevated in CO2 treated tumors compared to control. We also observed an enrichment of cytochrome c in the cytoplasmic fraction and Bax protein in the mitochondrial fraction of CO2 treated tumors, highlighting the involvement of mitochondria in apoptosis. These data indicate that transcutaneous application of CO2 may represent a novel therapeutic tool in the treatment of human MFH.

Transcutaneous Application of Carbon Dioxide (CO2) Enhances Chemosensitivity by Reducing Hypoxic Conditions in Human MalignantFibrous Histiocytoma

Background: Tumor hypoxia is a common feature of various human malignancies. Hypoxia contributes to tumor progression, and is a major cause of tumor resistance to chemotherapy. Hypoxia-inducible factor (HIF)-1 is a key transcription factor in hypoxic responses, and regulates the transcription of genes that are involved in crucial aspects of cancer biology, including angiogenesis, cell survival, and invasion. We previously demonstrated that transcutaneous application of carbon dioxide (CO2) induced oxygenation in the treated tissue in vivo, therefore, we hypothesized that transcutaneous CO2 exposure could enhance the chemosensitivity by reducing hypoxia in a tumor tissue. The aim of this study was to examine the effect of oxygenation by transcutaneous application of CO2 on the therapeutic efficacy of doxorubicin (DOX) to treat human malignant fibrous histiocytoma (MFH) in vivo. Methods: In this study, we utilized a murine model of human MFH, and mice were randomly divided into four groups: control, CO2, DOX and combination (CO2 + DOX) treatment groups to examine the effect of transcutaneous application of CO2 on the hypoxic condition, and to assess the therapeutic effect of combination therapy using transcutaneous CO2and DOX treatment in vivo. Results: Transcutaneous application of CO2 treatment decreased HIF-1α expression in human MFH tumor tissues, suggesting that our transcutaneous CO2 treatment reduced the hypoxic conditions. Furthermore, transcutaneous CO2 treatment alone had an antitumoral effect, and increased the chemotherapeutic effect of DOX on MFH tumor growth in vivo, with no observable effects on body weight. Conclusions: Our findings in this study strongly indicate that our transcutaneous CO2 system has antitumor effects and can enhance the chemosensitivity of tumor cells by reducing the local hypoxic conditions.

‘Decoy’ and ‘non-decoy’ functions of DcR3 promote malignant potential in human malignant fibrous histiocytoma cells

Decoy receptor 3 (DcR3) is a soluble secreted protein that belongs to the tumor necrosis factor receptor (TNFR) superfamily. DcR3 inhibits the Fas ligand (FasL)/Fas apoptotic pathway by binding to FasL, competitively with Fas receptor. Previous studies have reported that overexpression of DcR3 has been detected in various human malignancies and that DcR3 functions as a ‘decoy’ for FasL to inhibit FasL-induced apoptosis. In addition, recent studies have revealed that DcR3 has ‘non-decoy’ functions to promote tumor cell migration and invasion, suggesting that DcR3 may play important roles in tumor progression by decoy and non-decoy functions. We have previously reported that overexpression of DcR3 was observed in human malignant fibrous histiocytoma (MFH), however, the roles of DcR3 in MFH have not been studied. In the present study, to elucidate the roles of DcR3 in tumor progression of MFH, we examined the effects of DcR3 inhibition on cell apoptosis, migration and invasion in human MFH cells. siRNA knockdown of DcR3 enhanced the FasL-induced apoptotic activity and significantly decreased cell migration and invasion with a decrease in the activation of phosphatidylinositol 3 kinase (PI3K)/Akt and matrix metalloproteinase (MMP)-2. The findings in this study strongly suggest that DcR3 plays important roles in tumor progression of human MFH by decoy as well as non-decoy functions and that DcR3 may serve as a potent therapeutic target for human MFH.

Usefulness of MRI for diagnosis of painful snapping elbow.

Painful snapping of the elbows is rare. We report on a 12-year-old boy with a painful snap in both elbows. High-resolution magnetic resonance imaging of the elbow using microscopy coils detected a synovial fold interposed in each humeroradial joint and was very helpful in establishing the cause of symptoms. Resection of the synovial folds was performed with subsequent relief of symptoms.

Protein kinase Cδ in tumorigenesis of human malignant fibrous histiocytoma

Protein kinase Cδ (PKCδ), an isoform of PKC, has been shown to act as a critical mediator of tumor progression and apoptosis; however, its role in musculoskeletal tumors is still unknown. In the current study, we examined the expression of PKCδ in human musculoskeletal tumor tissue samples, and investigated the effects of siRNA downregulation of PKCδ on human malignant fibrous histiocytoma (MFH) cell proliferation, migration, and apoptosis, to elucidate its functional roles in musculoskeletal tumorigenesis. Of note, real-time PCR analysis revealed that mRNA expression of PKCδ in high-grade musculoskeletal MFH tumors was significantly lower than that in benign schwannomas. siRNA downregulation of PKCδ significantly increased human MFH cell proliferation and migration, and markedly suppressed apoptosis. These findings suggest that PKCδ has a negative effect on tumorigenesis and/or acts as a pro-apoptotic kinase in human MFH cells. The data presented here could be applied in the development of new therapeutic avenues, with the elevation of PKCδ expression being one potential strategy to prevent MFH progression. Thus, PKCδ may be a potent therapeutic target for human MFH.

AICAR induces mitochondrial apoptosis in human osteosarcoma cells through an AMPK-dependent pathway

The AMP-activated protein kinase (AMPK) activator 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) modulates cellular energy metabolism, and promotes mitochondrial proliferation and apoptosis. Previous studies have shown that AICAR has anticancer effects in various cancers, however the roles of AMPK and/or the effects of AICAR on osteosarcoma have not been reported. In the present study, we evaluated the effects of AICAR on tumor growth and mitochondrial apoptosis in human osteosarcoma both in vitro and in vivo. For in vitro experiments, two human osteosarcoma cell lines, MG63 and KHOS, were treated with AICAR, and the effects of AICAR on cell growth and mitochondrial apoptosis were assessed by WST assays, TUNEL staining, and immunoblot analyses. In vivo, human osteosarcoma-bearing mice were treated with AICAR, and the mitochondrial proliferation and apoptotic activity in treated tumors were assessed. In vitro experiments revealed that AICAR activated AMPK, inhibited cell growth, and induced mitochondrial apoptosis in both osteosarcoma cell lines. In vivo, AICAR significantly reduced osteosarcoma growth without apparent body weight loss and AICAR increased both mitochondrial proliferation and apoptotic activity in treated tumor tissues. AICAR showed anticancer effects in osteosarcoma cells through an AMPK-dependent peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α)/mitochondrial transcription factor A (TFAM)/mitochondrial pathway. The findings in this study strongly suggest that AICAR could be considered as a potent therapeutic agent for the treatment of human osteosarcoma.

Boron neutron capture therapy (BNCT) as a new approach for clear cell sarcoma (CCS) treatment: Trial using a lung metastasis model of CCS.

Clear cell sarcoma (CCS) is a rare malignant tumor with a poor prognosis. In the present study, we established a lung metastasis animal model of CCS and investigated the therapeutic effect of boron neutron capture therapy (BNCT) using p-borono-L-phenylalanine (L-BPA). Biodistribution data revealed tumor-selective accumulation of (10)B. Unlike conventional gamma-ray irradiation, BNCT significantly suppressed tumor growth without damaging normal tissues, suggesting that it may be a potential new therapeutic option to treat CCS lung metastases.